1. Technical Field
The present invention relates to a sucrose phosphate synthase from Citrus including its isoform, and to DNA encoding the enzyme.
2. Earlier Technology
Sucrose is a transport form of the photoassimilate in most plants. The sucrose in mature leaves (source) is mostly transported by the phloem to the plant organs (sink) that are net consumers of the photo-assimilate. A key enzyme of sucrose synthesis pathway, sucrose-phosphate synthase (EC 2.4.1.14) (hereinafter referred to as "SPS"), catalyzes the following reaction:
Fructose 6-phosphate+UDP glucose.fwdarw.Sucrose 6-phosphate+UDP Sucrose 6-phosphate is converted to sucrose by sucrose phosphatase.
Considerable interest has focused on the role of SPS in regulation of sucrose synthesis in source leaves (Kerr, P. S. and Huber, S. C. (1987) Planta 170:197-204; and Echeverria, E. and Burns, J. K. (1989) Plant Physiol. 90:530-533). SPS activity itself has been found in many plants, for example cucurbits (Lingle, S. E. and Dunlap, J. R. (1987) Plant Physiol. 84:386-389; Hubbard, N. L. et al. (1989) Plant Physiol. 91:1527-1534; and Burger, Y. and Schaffer, A. A. (1991) Sucrose metabolism in mature fruit peduncles of Cucumis melo and Cucumis sativus. In:Recent advances in phloemtransport and assimilate partitioning, pp. 244-247, Bonnemain, J. L. et al. eds. ouest Editions, Nantes, France), peach (Hubbard, N. L. et al. (1991) Physiol. Plant 82:191-196), pear (Moriguchi, T. et al. (1992) J. Am. Soc. Hortic. Sci. 117:247-278), and celery (Stoop, J. M. H. and Pharr, D. M. (1994) J. Am. Soc. Hortic. Sci. 119:237-242), sugar beet (Fieuw, S. and Willenbrink, J. (1987) J. Plant Physiol. 131:153-162), sugar cane (Wendler, R. et al. (1990) Planta 183:31-39; and Goldner, W. et al. (1991) Plant Sci. 73:143-147), sucrose-accumulating Lycopersicon spp. (Miron, D. and Schaffer, A. A. (1991) Humb. And Bonpl. Plant Physiol. 95:623-627), Dali, N. et al. (1992) Plant Physiol. 99:434-438; and Stommel, J. R. (1992) Plant Physiol. 99:324-328), rice (Smyth, D. A. and Prescott, H. E. (1989) Plant Physiol. 89:893-896), strawberry (Hubbard, N. L. et al. (1991) Physiol. Plant 82:191-196), and citrus (Lowell, C. A. et al. (1989) Plant Physiol. 90:1394-1402; and Echeverria, E. (1992) Plant Sci. 85:125-129).
To investigate the enzymatic function of SPS on sucrose biosynthesis, SPS has been purified to near homogeneity from spinach (Salvucci, M.E. et al. (1990) Arch. Biochem. Biophys. 281:212-218), wheat (Salerno, G. L. et al. (1991) Physiol. Plant 81:541-547), and maize (Bruneau, J. -M. et al. (1991) Plant Physiol. 96:473-478). SPS is an allosteric enzyme which is activated by binding of the substrate-similar glucose-6-phosphate and inhibited by P.sub.i at the allosteric site (Doehlert, D. C. and Huber, S. C. (1983) Plant Physiol. 73:989-994). In addition, the activity of SPS is regulated by protein phosphorylation (Huber, J. L. A. et al. (1989) Arch. Biochem. Biophys. 270:681-690; Siegl, G. et al. (1990) FEBS Letters 270:198-202; and Huber, S. C. and Huber J. L. (1991) Plant Cell Physiol. 32:319-326). Recently, the function and structure of SPS have also been studied at the molecular level in maize (Worrell, A. C. et al. (1991) Plant Cell 3:1121-1130), spinach (Klein, R. R. et al. (1993) Planta 190:498-510), and sugar beet (Hesse, H. et al. (1995) Mol. Gen. Genet. 247:515-520).
In citrus, the sucrose accumulation is one of the very important events in fruit development. Phloem-free juice sacs at the middle stage of fruit development showed higher SPS activity than the adjacent transport tissues, vascular nodules and segment epidermis (Lowell, C. A. et al. (1989) Plant Physiol. 90:1394-1402). However, analysis of the function and expression of SPS at the molecular level has been quite limited in Citrus.
An object of this invention is to clone cDNA for SPS from Citrus and characterize it at the molecular level.
Another object of the invention is to provide an SPS from Citrus.